I believe that as the surface area increases, it will mean that more oxygen will be produced, until all the hydrogen peroxide has been broken down and the enzymes can’t break any more down, the amount of oxygen produced will decrease until there isn’t any being produced. Or, there aren’t enough catalase enzymes in a certain surface area, meaning that ALL the enzymes will be working, in a sufficient amount of hydrogen peroxide, so the amount of hydrogen peroxide given off will plateau, because it is the maximum amount of oxygen that can be given off by using that number of enzymes until the amount of hydrogen peroxide to be broken down by the enzyme runs out and the amount of oxygen produced will decrease. This will be due to an increase in the number of active sites available to breakdown the hydrogen peroxide into water and oxygen. The higher the number of available active sites, means more hydrogen peroxide molecules can be broken down.
To make sure that my experiment runs as fairly as accurately as possible I have to find out what factors affect enzymes. As temperature increases, molecules move faster (kinetic energy theory). In an enzyme catalysed reaction, such as the break down of hydrogen peroxide, this increases the rate at which the enzyme and substrate molecules meet, and therefore the rate at which the products are formed. As the temperature continues to rise, however, the hydrogen and ionic bonds, which hold the enzyme molecules in shape, are broken. If the molecular structure is disrupted, the enzyme ceases to function as the active site is no longer the correct shape for the substrate and the enzyme can’t break down the substrate. To make sure that the heat from my hands didn’t affect the results I tried to make sure that I only touched the top of the boiling tubes and not the bottom part where the liquid was. Any change in the pH of a solution will effect how the enzyme works. By changing the pH of the solution which the enzyme is in, it disrupts the size and shape of the active site and the enzyme can’t work and is denatured. Enzymes have an optimum pH at which they can work in, when this changes the output of the enzyme also changes. In my experiment I made sure that nothing which could affect the pH could get into the boiling tube which the enzyme was in; to do this I washed the boiling tubes with water and dried them thoroughly afterwards. If there is an excess of enzyme molecules, an increase in the substrate concentration, produces a proportional increase in the rate of reaction. If there are sufficient substrate molecules to be broken down by the enzyme, the rate of reaction is unaffected by further increase in substrate concentration as the enzymes are unable to break down the greater quantity of substrate. Inhibitors are other substrates which compete with the original substrate (hydrogen peroxide) for the active sites, this is known as competitive inhibition. Another substrate could also attach itself to the enzyme altering the size and shape, and therefore not allowing it to break down the original substrate, non-competitive inhibition. Other types of inhibitors include, irreversible inhibition, this is when a substrate enters the active site and can’t be removed, making the enzyme useless. Enzyme concentration, as long as there is an excess substrate, an increase in enzyme concentration will lead to a corresponding increase in rate of reaction. Where the substrate is in short supply (i.e. it is limiting) an increase in enzyme concentration will have no effect. I varied the enzyme concentration by altering the number of equal sized discs of potato that contain the catalase, in the reaction.
By finding out the above information, I now know what factors will effect my experiment, I have had to design an experiment which takes into account the above factors.
If I changed any of the above factors whilst carrying out the investigation, it would result in a change with the final results and the experiment wouldn’t show any kind of relationships and the experiment would have been a waste of time.
By using the formula 2 H2O2 >-Catalase-> 2 H20 + O2 I can see that the hydrogen peroxide will be broken down into oxygen and water. In order to measure the reaction I will have to measure the amount of oxygen or water produced. I have found that oxygen is an easier product to measure than water.
I think, as the surface area increases, then the time in which the given amount of oxygen to be produced will decrease and the graph should look something like the following graph.
From the graph I have predicted that the time taken for the given amount of oxygen to be produced will decrease up to a certain point. When this point is reached I should expect to see a slowing down in the time taken as all the substrate has been used up. This means, as I said earlier, that no more oxygen can be produced as there isn’t anything to make the oxygen from.
To measure the oxygen, I had to design an experiment which could capture the oxygen produced, to do this I created the following experiment.Apparatus
- Tub filled with water
- Clamp
- Two clamp stands
- Bung
- Tubing
- Boiling tube
- Inverted measuring cylinder
- 2 molar Hydrogen peroxide
- Potato
- Size 6 potato gorger
- Razor blade
- An old tile
- Ruler
- Stopwatch
- Measuring Cylinder
I used the above equipment for accuracy, as you can see in the diagram below, the clamp stands are placed in a position so they can hold their tubes over the water in a particular position. By keeping these clamp stands in the same place for every experiment, it will lower the chance that anything could differ with the experiments, raising the accuracy rates.
I had to find a way of collecting the amount of oxygen produced, this was very hard to measure, but I found that measuring how much water could be displaced was a better method because it wouldn’t involve any machines and wouldn’t become complicated. If any mistakes were made, then it would be through human error. By creating many tables of results, I would be able to find out which results were right, and which ones weren’t. To follow this method, it involved oxygen to be collected in a measuring cylinder filled with water to see how much water was displaced.
I used the Size 6 gorger and kept this constant all the way through my experiment otherwise the number of active sites available would be different allowing different amounts of oxygen and water to be collected.
I used the highest molar solution, 2.0, because it means that most of the active sites on the potato would be taken up and wouldn’t be inactive because there wasn’t enough substrate to be broken down. When all the substrate has been broken down, the amount of oxygen and water produced, should plateau, this is why I am expecting my results to be closer together towards the end of the experiment.
I created the experiment in the following way…
Method
- To start the experiment, take the size 6 potato gorger and gouge a few pieces of potato.
- Take the gouged pieces of potato and cut them, by using the ruler and a razor blade, into lengths of 2 cm’s, but make sure the end of the potato isn’t used and is discarded. This is because the skin of the potato may have different properties to the actual inner pieces of the potato and would change the results from the experiment.
- By using a ruler cut one of the potato pieces 20mm from one of the ends.
Potato.
Cut like this. Cut
- Place all of the potato into the boiling tube.
- Measure out 10ml of 2.0 molar hydrogen peroxide. By using a strength of 2.0 molar hydrogen peroxide it will allow most of the catalase’s active sites to be used, resulting in the quickest reaction possible, making it easier and quicker for me to carry out all of the intended experiments.
- Place the clamp on the inverted measuring cylinder then fill it with water. To do this turn it upside down in the water and then clamp it into place but not allowing the base to rise above the water level.
- Make sure the stop watch is set to 0 and make sure that the tubing is inside the inverted measuring cylinder.
- Pour the hydrogen peroxide into the boiling tube, push the bung on top, start the stop watch and see how long it takes for enough oxygen to be produced for the water level in the inverted measuring cylinder to be at 3.0 ml
Carry out this test 9 more times, but for each different test make an extra cut (Part 3) 20mm further down the potato. Record the results in a table and repeat these set of ten experiments a further 3 more times. After each individual experiment, wash out the boiling tube and use a different piece of potato. As this test involves the use of hydrogen peroxide, it is vital that goggles are worn, and any loose clothing is either removed or made safe. Also, when cutting the potato, the potato is wet, this makes it hard to cut, be careful when using the razor blade.
These are my recorded results:
Time in seconds in which it took 3cm 3 of oxygen to be produced:
These surface areas were calculated by using the following formula…2πr(r+h)
For Example, for my first experiment I didn’t have any cuts… so the surface area was found out by doing these calculations:
2πr = 2 x π x 50
= 314.1592654mm2
This then has to be multiplied by (r+h)
50 + 200 = 250
314.1592654 x 250 = 78539.81634mm2
By using Spearman’s Rank theory, I found out that as the surface area increase, the reaction time decreased. Spearman’s Rank shows that the relationship between the two sets of data is perfect correlation.
Perfect correlation shows that the two sets of data are related in the strongest possible way. This proves that as the surface area increases, the average time taken, decreases!
The following table enforces the fact that as the surface area increases, the time taken for the 3.0cm3 of oxygen to be produced also declines.
Difference of results during the experiment:
When I look at my graphs, I can see that there are a few anomalies. I believe this is due to a few factors, which I will mention in my evaluation. From looking at my results i found that as the surface area doubles in size, the time taken for the 3.0cm2 to be produced halved.
Comparison between two results:
From this, I can expect, assuming that the conditions are ideal, that is I were to double the surface area, the time taken to produce the 3.0ml of oxygen should half. This is mainly due to the same reason which I mentioned in my preliminary work, by doubling the surface area it allows double the amount of catalase molecules to react with the hydrogen peroxide. Which means double the amount of oxygen can be produced.
Evaluation
As I stated in my planning part of this investigation, I predicted that, “hydrogen peroxide will breakdown to oxygen and water in the presence of Catalase. The reaction will increase with increasing enzyme concentration when molecules of hydrogen peroxide are freely available. However, when molecules of the substrate are in short supply, the increase in rate of reaction is limited and will have little effect.” I believe that my hypothesis is correct. Although my readings did show that towards the end of the experiment, the amount of oxygen being produced was slowing down, I believe this was because there was a lack of the substrate available to be broken down, thus resulting in the enzyme efficiency slowing down.
I felt that I used the best available strength molar of hydrogen peroxide because if I had used a 1.0 strength there would have been less of the substrate to be broken down and the amount of oxygen produced would have levelled off a lot earlier than it did. I wouldn’t have wanted this due to the fact that I needed ideal conditions to work out if there is a relationship between the amount of oxygen produced and the surface area.
Whilst carrying out my investigation I was using an inverted cylinder to measure the amount of oxygen produced. The cylinder had marks every 0.5cm3, this made it very hard to estimate when the 3.0cm3 of oxygen was produced. This would have meant that I wouldn’t know exactly when the 30.cm3 of oxygen was produced affecting the results in my table. Although this was a major mistake, it did happen to all the results that I collected and, in general, it wouldn’t change the readings completely.
Another part of human error occurred when I was cutting the potato. I realised that my ruler was in mm, this meant that there would be an error in length of the potato when trying to cut it to 2.0 cm. I have also realised that whilst cutting the potato I was cutting it at different angles. Ideally I would have liked to cut them at a right angle, otherwise the amount of surface area would be different. Again, this mistake happened to all of my results and I shouldn’t think that the overall results were affected greatly. I think a good way of solving the above two issues is to use a liquid form of catalase. Although this would be very accurate, and I could measure the amount of catalase is reacting with the hydrogen peroxide. However, I wouldn’t be finding out the effect of increasing amounts of surface area of potato, reacting with hydrogen peroxide, unless I knew how much catalase is produced per mm2 of potato.
When I measured out the amount of hydrogen peroxide which I should’ve been using, I didn’t use a pipette to pour the solution into the measuring cylinder, this could have meant that there were many variations of hydrogen peroxide reacting with the catalase.
Due to the size of this investigation, I had to carry out the experiments over the course of a few days, each day I had to set up the apparatus again. Even though I tried to place the apparatus in the same place every time, the position would have changed, allowing different readings. Ideally, I should have carried out the investigation during one day and using the same apparatus. By carrying out the experiment over a few days, I had to use different potatoes, the different potatoes would have probably been containing different amounts of catalase, and therefore changing my results even further.
Another mistake which I found was when I placed the potato into the boiling tube and I then poured the hydrogen peroxide in, I couldn’t have placed the bung on in the boiling tube and started the stopwatch as soon as possible. There were bound to be a few seconds difference.
Although I think the experiment isn’t a very accurate one in terms of the number of variables affecting the results, I feel that the errors were small and couldn’t have made a major difference to the overall readings. Any of the errors which I mentioned above all occurred to each test. If all the tests were affected in the same way, then the results couldn’t have changed much.
Whilst looking at my graphs, I noticed a couple of anomalies, they have been indicated on my graphs. I believe that these anomalies were caused by some of the errors which I mentioned above, although they wouldn’t have made a major difference on their own, I feel that some of the errors would have happened to some test and maybe no errors happened to the others. This could also work both ways, maybe all the errors above happened to these anomalies, each error may have not affected the test much, but when all the errors occur then the total error could be massive and this is probably what happened with my anomalies.
I think the best way to improve this experiment is to use a gas syringe instead of using the inverted measuring cylinder syringe. By using a gas syringe, it means that there won’t need to be any water involved as the whole experiment is enclosed. This will reduce the amount of variables in the experiment, making it a lot more accurate. The gas syringe will be pushed up from the extra pressure from the releasing oxygen. Making it an easier way to capture the correct amount of oxygen.
I conclude, from my experiment, that my hypothesis is correct and as surface area increases, so does the rate of reaction between catalase and hydrogen peroxide.